成纤维细胞在不同形貌钛合金表面上的生物学行为

发布时间：2018-06-06 16:31

本文选题：电解蚀刻 + 钛合金　；参考：《吉林大学》2014年硕士论文

【摘要】：目的：种植体的成功标准除了具有良好的骨结合,还要求形成良好的种植体周软组织封闭。影响种植体周软组织封闭的重要决定因素为种植体颈部的表面形貌。本实验旨在比较通过机械加工法(M)、直流电解蚀刻法(DC)、交流电解蚀刻法(AC)三种不同的方法处理钛合金试件得到的三种不同的表面形貌对人牙龈成纤维细胞的附着的影响，寻找出一种有利于成纤维细胞生长以及软组织附着的理想表面，为今后种植体颈部设计提供指导。方法：通过机械加工法（M）、直流电解蚀刻法(DC)、交流电解蚀刻法(AC)成形三种不同钛合金表面形貌，使用扫描电子显微镜观察其表面微观形态，将其与成纤维细胞共同培养，吖啶橙染色法检测生物相容性；MTT法检测表面细胞的增殖情况；扫描电镜观察表面细胞的形状及排列。结果：通过机械加工(M)、直流电解蚀刻(DC)、交流电解蚀刻(AC)三种处理方法分别得到了较光滑的表面、微米-亚微米孔表面、微米-亚微米-纳米结构表面。通过吖啶橙染色法显示成纤维细胞在三种表面形貌的钛片上均具有良好的细胞活性，核仁清晰，胞浆饱满，提示三种表面形貌的钛片均具有良好生物相容性。MTT结果显示，三种表面形貌的钛片吸光度值变化趋势基本一致，均呈递增趋势。光滑表面组与微米-亚微米表面组和微米-亚微米-纳米组表面吸光度值有统计学差异。在第3-5天，微米-亚微米-纳米组表面吸光度值高于微米-亚微米表面，提示成纤维细胞在微米-亚微米-纳米表面的增殖情况最优。将成纤维细胞与三组钛片共同培养的第3天，扫描电镜观察可见，各组钛件表面胞外基质较少，可见明显未被基质覆盖的沟、凹孔形态。M组钛片低倍镜下可见细胞扁平，呈圆形，近似平行排列，细胞长轴与沟纹方向一致，伸展较差，高倍镜下可见细胞伸出许多微绒毛，伪足较少且短；DC组低倍镜下可见细胞附着于微米孔内，伸展较好；高倍镜下可见细胞丰满，细胞表面微绒毛及突起较少，伸出的伪足较粗较长，锚固于微米孔壁上；AC组低倍镜下可见细胞附着于微米孔内，形态更加伸展，高倍镜下可见细胞丰满，细胞表面有许多突起及微绒毛，伸出的伪足更宽更长，可成带状，相邻细胞的伪足可发生融合。将钛片与成纤维细胞共同培养的第7天，扫描电镜观察可见，各组钛件表面胞外基质较第3天明显增多。M组低倍镜下可见材料表面沟纹被细胞外基质大面积覆盖，细胞呈圆形，伸展较差；高倍镜下，细胞伸出的微绒毛紧密的贴在材料表面，细胞伸出的伪足较第3天增长增多，相邻细胞的伪足可相融合。DC组低倍镜下可见微米孔被大量细胞外基质覆盖；高倍镜下可见细胞完全展开，呈扁平片状，少量细胞呈球形，可能为基质或细胞表面上新分裂的细胞，细胞伸出许多伪足和微绒毛，紧紧附着于微米孔的孔壁上。AC组低倍镜下可见细胞分泌的基质大面积覆盖在材料表面，高倍镜下可见细胞形态更加扁平不规则，细胞胞体位于微米孔内，向邻近微米孔壁伸出大量伪足及绒毛，伪足可相互交织成网状；细胞牢固的固定于微米孔中，可见长梭形及球形新生细胞，有的细胞伸出细长的伪足，呈悬空状，可伸入到相隔较远的纳米孔内部。结论：通过交流电解蚀刻法形成的兼具微米-亚微米-纳米三级孔结构的钛合金表面具有良好的生物相容性，能够促进成纤维细胞的附着，，增殖及细胞基质的分泌。通过该方法处理出的种植体颈部表面是一种较优秀的表面，可为以后的相应体内实验及种植体表面处理提供指导。
[Abstract]:Objective : To study the effect of three different surface morphologies on the adhesion of the implant neck with three different methods : mechanical machining ( M ) , direct current etching ( DC ) , AC electrolytic etching ( AC ) , and to find an ideal surface for the growth of fibroblasts and the attachment of soft tissues , which will provide guidance for the neck design of implants .

Methods : The surface morphology of three different titanium alloys was formed by mechanical machining ( M ) , direct current etching ( DC ) and alternating current electrolytic etching ( AC ) . Scanning electron microscope ( SEM ) was used to observe its surface morphology , which was co - cultured with fibroblasts , and acridine orange staining was used to detect biocompatibility ;
MTT assay was used to detect the proliferation of surface cells .
Scanning electron microscope was used to observe the shape and arrangement of surface cells .

Results : The results showed that three kinds of surface morphology of fibroblasts were obtained by mechanical processing ( M ) , direct current etching ( DC ) and alternating current electrolytic etching ( AC ) .
In DC group , the visible cells were attached to the micro - pores and extended well .
Under the microscope , the visible cells are plump , the surface microvilli and the protrusion of the cells are less , the extended pseudopod is thicker and longer , and is anchored on the wall of the micron pore ;
Under the low - magnification microscope , the visible cells were attached to the micrometer pores , the morphology was more extended , the cells under the microscope were full , the surface of the cells had a lot of protrusions and microvilli , and the surface of the cells was wider and longer . The outer matrix of the surface of the titanium pieces was obviously increased on the 7th day . The surface of the surface of the titanium pieces was covered with large area of the extracellular matrix , and the cells were round and stretched .
Under the microscope , the microvilli extending from the cells were closely attached to the surface of the material , and the pseudopods of the cells extended more than the third day , and the pseudopodium of adjacent cells could be fused .
Under high magnification , the visible cells are completely expanded , flattened , and small cells are spherical , which may be newly divided cells on the surface of the matrix or cells . The cells extend out of many pseudopods and microvilli and adhere tightly to the pore walls of micron pores . The cells secreted by the cells under the low magnification of the AC group are covered on the surface of the material .
The cells are firmly fixed in the micro - pores , and can be found to have long fusiform and globular new cells , and some cells extend out of the slender pseudopod , and are in a suspended shape , and can extend into the nano - pores which are spaced farther apart .

Conclusion : The surface of titanium alloy with micron - submicron - nanometer three - stage pore structure formed by the method of AC electrolytic etching has good biocompatibility , which can promote the adhesion , proliferation and secretion of cell matrix . The surface of implant neck treated by this method is an excellent surface , which can provide guidance for the subsequent in vivo experiment and implant surface treatment .
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R783.6

【参考文献】